Επίδραση διαφόρων παραγόντων στη υάλωση, βλαστογένεση και ριζοβολία της ελιάς (Olea europaea L.) in vitro

Abstract

The present study consists of three experiments concerning the effect of various parameters on the micropropagation stages and the problem of hypehydricity of olive explants (Olea europaea L.). In the 1st Experiment the in vitro establishment of different Greek and foreign olive cultivars (‘Amfissis’, ‘Galatistas’, ‘Kothreiki’, ‘Koroneiki’, ‘Lianolia Kerkiras’, ‘Ladolia Patron’, ‘Chondrolia Chalkidikis’, ‘Arbequina’, ‘FS-17’, ‘Manzanillo’ and ‘Picual’) was tested. The initiation of micropropagation using young or mature shoots from adult field grown trees was difficult due to high contamination, hyperhydricity and strong oxidation of tissues. On the other hand, explants derived from greenhouse growing stock plants were a better plant source for the in vitro culture of ‘Chondrolia Chalkidikis’ and ‘Arbequina’. However, the in vitro establishment of ‘Arbequina’ was not successful since the axillary buds did not develop new shoots. The explants of ‘Chondrolia Chalkidikis’ were disinfected by treating them with a solution of commercial bleach (20%) for 10 min. The removal of the nodal leaves enhanced the development of axillary shoots. Moreover, the explants of ‘Chondrolia Chalkidikis’ presented hyperhydricity symptoms, thus limiting the formation and rooting of microshoots in vitro. The hyperhydrated shoots were ‘wet’, while the leaves were thicker, dark green, fragile and curled. In the 2nd Experiment trials on ‘Chondrolia Chalkidikis’ were made in order to enhance the production of new shoots of good quality. Olive Medium (OM) stimulated the development of axillary shoots, without any severe abnormalities. Doubling the concentration of the macronutrients of Woody Plant Medium (WPM) promoted the shoot length. The pH of the OM was adjusted to 5.8 before autoclaving. Increasing the pH of the medium did not inhibit hyperhydricity. Among the cytokinins tested, zeatin (2 mg l-1) stimulated the production of more and longer shoots while thidiazuron (TDZ) inhibited shoot length growth. Both cytokinins presented fewer hyperhydricity symptoms. Benzyladenine (BA) also led to the formation of a satisfactory number of new shoots. Moreover, it increased the percentage of ‘wet’ shoots. Hyperhydricity was limited after dipping the base of the explants in a BA solution (100 mg l-1) for 1 min and subsequently transferring them onto a hormone-free medium. Moreover, this treatment was adequate to promote axillary shoot development. The simultaneous presence of BA and gibberellic acid (GA3) (2.0+2.0 mg l-1, respectively) in the OM induced less symptoms than did BA alone. Keeping the explants only for 2 days in the OM containing BA and then culturing them onto a medium with 2.0 mg l-1GA3 did not promote either hyperhydricity or shoot production. Expanding the exposure to 4 days also did not enhance the number of the shoots formed. Besides 2.0 mg l-1 BA, including the growth inhibitor dikegulac (100.5 μM) in the OM increased the number of new long shoots, with less hyperhydricity symptoms. Similarly, less severe hyperhydricity and enhanced shoot production was achieved after supplying the medium with 10 μM AgNO3. On the other hand, CoCl2 and salicylic acid did not give good results. Replacing agar with vermiculite enhanced the production of good quality shoots. Hyperhydricity was more intense in the semi-solid OM containing 6 g l-1. Increasing the concentration of agar reduced not only hyperhydricity but also shoot formation. Hyperhydrated shoot cultures did not recover after transferring them from the semisolid to the medium containing vermiculite. Keeping the olive shoot cultures under yellow or blue light tubes did not affect hyperhydricity but stimulated the number of the shoots produced. In the 3nd Experiment in vitro rooting of ‘Chondrolia Chalkidikis’ was achieved after establishing the microshoots onto ½OM containing 1 mg l-1 ### and after partially darkening the base of the shoots (the lower part of the test tubes was covered with black plastic tape). This treatment also increased the survival rate of the rooted olive plantlets during acclimatization.